The aim of this research is to elucidate the protein structural requirements for electron transfer and control in the mitochondrial electron transfer chain, and in the microsoma 1 NADPH cytochrome c (P450) reductase system. It is proposed that control of oxygen consumption at the level of cytochrome oxidase may involve a mechanism similar to that define for hemoglobin, in which changes in electrostatic interactions between subunits determine the affinity of oxygen binding at the heme. In the case of cytochrome oxidase, the strong ionic interactions with its substrate, cytochrome c, are likely to influence the conformation of the oxidase, as is the membrane potential gradient. The effects of the presence, absence and binding affinity of cytochrome c on respiratory control in coupled inner mitochondrial membranes will be studied. Steady-state, pre-steady state, and low-temperature kinetic analyses of the reactions of native and modified cytochromes c with cytochrome c oxidase will be carried out with a purified, mono disperse oxidase preparation, as well as with the membrane-bound form, in order to further clarify the mechanism and mechanics of the reaction. This will also require direct binding studies and an investigation of the kinetic effects of covalently cross-linking cytochrome c to purified and membrane-bound oxidase. The reaction of cytochrome c with microsomal NADPH-cytochrome c (P450) reductase will be investigated as a function of cytochrome c structure, ionic strength, presence of cytochrome P450, and the type of reductase preparation (protease-solubilized, detergent solubilized, and membrane-bound), in order to define the nature of the protein interactions involved.